[Proteolytic enzymes in human lymphocytic leukemia cells. I. Activity of dipeptidylaminopeptidase IV, plasminogen activator and cathepsins B and L in cells with different immunologic phenotype]. / Proteoliticheskie fermenty v leikoznykh limfoidnykh kletkakh cheloveka. I. Aktivnost' dipeptidilaminopeptidazy IV, aktivatora plazminogena i katepsinov B i L v kletkakh s raznym immunologicheskim fenotipom.

Immunological and biochemical study of lymphoid cells obtained from 20 patients with various forms of lymphoproliferative disorders has been carried out. It was found that different phenotypes of lymphoid cells at various stages of differentiation have different activity levels of dipeptidyl aminopeptidase IV (DAP IV), plasminogen activator (urokinase type) and cathepsins B + L. The highest proteinase activity values were found in the lysates of just those leukemic T-cells whose phenotypes corresponded to the initial stages of thymic differentiation or to activated T-cells. The 10 times lowered activity was found in the cell phenotypes of mature T-helpers and T-suppressors, and the activity of the both was at virtually the same level. In lymphoid cells of the B-lineage (from pre-B to mature B-lymphocytes) the proteinase activities did not differ essentially: they were 2 to 3 times lower than in the lymphoid progenitors. It was suggested that the regulated activity changes in some proteinases occur during differentiation along the T- or B-pathways. It is likely that the increases in DAP IV and cathepsins B + L activities are associated with the activation of mature lymphoid T- and B-cells. No direct correlation was found between the activity of either proteinase and the expression of any of the surface markers under study.

[Proteolytic enzymes in human lymphocytic leukemia cells. II. Comparative characteristics of proteolytic enzymes and their inhibitors in three differing types of lymphoid cells]. / Proteoliticheskie fermenty v leikoznykh limfoidnykh kletakh cheloveka. II. Sravnitel'naia kharakteristika proteoliticheskikh fermentov i ikh ingibitorov v trekh razlichaiushchikhsia tipakh limfoidnykh kletok.

A comparative study of proteolytic enzymes and their inhibitors in three human leukemic lymphoid cell populations has been carried out. The lysates of all lymphoid cells contained cathepsins D, B, L and H as well as serine trypsin-like proteinases, several aminopeptidases, dipeptidyl aminopeptidase IV and plasminogen activator (urokinase type). The activities of individual proteinases and their ratios in all cell types under study varied essentially, suggesting that lymphoid cells with different functions have different sets of proteolytic enzymes. FPLC chromatography of the lysates revealed the presence of inhibitors of cysteine and serine trypsin-like proteinases. The procedure for isolation of cathepsins D, B, L and H and of their inhibitors has been proposed and partially purified protein preparations obtained. Some properties of cathepsins B and L and those of their inhibitor have been examined.

Dipeptidylpeptidase IV in human leukemic B- and T-cells.

Dipeptidylpeptidase IV (DPP-IV) activity in lymphoid cells of patients with various lymphoproliferative diseases has been assayed. The enzyme activity was detected in lysates of all leukemic T- and B-cells studied. High DPP-IV activity levels (5-7 fold higher than those in mature Th and Ts) were found in T-cells with phenotypes corresponding either to the early thymic stages of maturation, or to activated T-lymphocytes, or to some atypic T-cells. In most leukemic B-cells with phenotypes of mature and late B-lymphocytes, the DPP-IV activity was lower than in mature T-cells; however, in some B-cells carrying the activation markers CD 25 and CD 30 the activity was rather high. High DPP-IV activity was seen in the majority of leukemic B-cells expressing CD 25 (87%) and CD 11c (82%) antigens. Experiments with intact cells have shown that DPP-IV is present in the plasma membrane of leukemic B- and T-cells studied. Thus DPP-IV cannot be considered as a marker of activated T-cells alone because a significant DPP-IV activity was also detected in a number of activated leukemic B-cells and in early T-cells.

[Protein kinase activity in lymphoid cells in various forms of lymphoproliferative disorders]. / Aktivnost' proteinaz v limfoidnykh kletkakh pri raznykh formakh limfoproliferativnykh zabolevanii.

Activities of dipeptidyl-aminopeptidase IV, urokinase-like plasminogen activator, cathepsins B and L were studied in lymphoid cells of patients with various forms of lymphoproliferative disorders. Activity of the enzymes studied was found in all the T- and B-cell, although rate and ratio of the enzymatic activity were dissimilar in various cell types. The highest rate of activity exhibited cells at early stages of maturation obtained from patients with acute lymphoblastic leukemia, while low level of the proteinase activity was detected in cells of patients with chronic lymphoid leukemia, non-Hodgkin lymphoma, hairy cell leukemia and Sezary disease, corresponding to mature T- and B-subpopulations. As shown by analysis of the cells immunological phenotype and their proteolytic activity, the rate of lymphoid cells differentiation correlated with level of proteinases activity. Series of proteinases were firstly studied in human malignant lymphoid cells with known phenotype. The enzyme assay may be used in diagnosis and treatment of patients with lymphoproliferative disorders.

Activity of some proteinases and glycosidases in human leukemic lymphoid cells at various stages of differentiation.

Activities of some glycosidases and proteinases in human leukemic lymphoid cells at various stages of differentiation have been compared. It was found that cells with different immunological phenotypes gave different enzymic spectra. Glycosidases and proteinases in lymphoid cell precursors had higher activity level than the enzymes in mature T- and B- cells. In cells of B- lineage, all activities were lower than in common precursor of lymphoid cells. In T-cells at the earlier stages of thymic differentiation, activities of all proteinases and most of glycosidases were higher than in common precursor cells whereas in mature T-helpers and T-suppressors the activities were markedly lower. Most of hydrolases in mature T-cells were twice more active than the enzymes in mature B-cells. The opposite-directional changes in activities of some hydrolases at the earlier stages of differentiation of lymphoid cells along B- or T- cells pathways are suggested.

[Dipeptidyl aminopeptidase-IV in lymphocytes of patients with lymphoproliferative diseases]. / Dipeptidilaminopeptidaza-IV v limfotsitakh bol'nykh limfoproliferativnymi zabolevaniiami.

DAP-IV activity (Gly-Pro-MCA hydrolysis, pH 7.8) was found in lysates of peripheral blood lymphocytes of patients with T- and B-cell forms of malignant lymphoproliferative diseases. The highest DAP-IV activity was seen in the cells of patients with a rare variant of T-cell lymphocytic leukemia (T-CLL); these cells expressed simultaneously the antigens of T helpers and T suppressors (Th and Ts) (OKT4+ and OKT8+). The DAP-IV activity about ten times less was found in the pathological cells with a phenotype of mature Th (Sezary disease), as well as in the cells expressing antigens of both Ts and natural killers (a rare variant of T-CLL). The same activity was also found in Ts (T gamma-lymphocytosis). The data obtained show that the differences in DAP-IV expression are connected with the differentiation step rather than with the belonging to a particular subpopulation of T-cells. DAP-IV activity, which was somewhat lower than that of T-cells, was found in B-lymphocytes of patients with B-CLL, hair-cellular leukemia, and non-Hodgkin's lymphoma. No correlation of DAP-IV activity with the level of E-cellular differentiation was observed.

[Isolation of neutral proteinase of ribosomes (cathepsin R)]. / Vydelenie neitral'noi proteinazy ribosom (katepsina R).

A method for isolation of cathepsin R from rat liver ribosomes allowing for a 264-fold increase of specific activity is described. The purification procedure includes enzyme extraction from ribosomes with 2-4 M LiCl and two-step affinity chromatography on Sepharose with immobilized soy bean trypsin inhibitor and trypsin-Sepharose.

[Inhibitors of cathepsin R (ribosomal proteinase). Polyamines as natural inhibitors of the proteinase]. / Ingibitory katepsina R (proteinazy ribosom) Poliaminy-estestvennye ingibitory proteinazy.

The effects of potential inhibitors on the activity of neutral ribosomal proteinase--cathepsis R--were studied. It was found that cathepsin R belongs to the group of serine enzymes. The polyamines spermine and spermidine, which are inherently present in the ribosomes, are natural reversible inhibitors of cathepsin R. Upon separation of the enzyme from the inhibitors the proteinase displays a high activity. The effects of polyamines on the proteinase activity may either be direct or mediated via RNA. The enzyme activity can also be controlled by amino acids. Approximately 2/3 of cathepsis R were found in a latent state.

[Specificity of neutral ribosomal protease]. / O substratnoi spetsifichnosti proteinazy ribosom.

The specificity of the ribosomal proteinase was studied using a variety of substrates: nascend peptides, ribosomal proteins, B-chain of insulin and some synthetic peptides (the heptapeptide Gly-Phe-Phe-Tyr-Thr-Pro-Lys; the hexapeptide Gly-Phe-Leu-Gly-Phe-Leu and the CBZ-hexapeptide; the tripeptide Phe-His-Leu and the CBZ-tripeptide). It was shown that in the heptapeptide tested the enzyme cleaved most rapidly the Phe-Tyr bond; the Phe-Phe bond was cleaved less rapidly. The rest of the peptide bonds in the heptapeptide were also cleaved and by the end of the incubation the cleavage was complete. All the other substrates tested also underwent complete degradation. The data obtained allowed us to classify the enzyme with the group of endopeptidases with broad specificity of action. The possibility cannot be excluded that the broad specificity observed is due to the presence of more than one enzyme on the polyribosomes.

[On the localization of ribosomal proteinase]. / O lokalizatsii neitral'noi proteinazy na ribosomakh.

The localization of proteolytic activity in rat liver ribosomes was studied. 40S and 60S subparticles were obtained upon dissociation of polyribosomes by puromycin. It was shown that the proteolytic activity is associated with small subparticles or with proteins removed from the latter.

Presence of a proteinase in polyribosomes of rat liver.

The presence of a proteinase in the composition of the ribosomal proteins of rat liver is demonstrated. The enzyme possesses optimal activity in the zone of pH 7.0-7.2. Soybean trypsin inhibitor and 1-chloro-4-phenyl-3-tosylamido-2-butanone inhibit the enzyme by 50-60%.

On the proteolytic acitivity of ribosomes. / O proteoliticheskoi aktivnosti pibosom

The presence of a proteinase on polysomes, isolated from rat liver has been demonstrated. The proteinase was not removed from polysomes upon treatment of the latter with 0,5 M ammonium chloride. The pH optimum of the enzymes is at pH 7,0.